Over past decades, the prior art has offered several types of rapid diagnostic testing techniques which use a body fluid such as whole blood, serum, plasma, urine, spinal fluid, amniotic fluid, mucous, saliva, and the like for detecting the presence of infection or other conditions such as cancer, pregnancy, abused drugs and cardiovascular disorders such as acute myocardial infarction (AMI). Such tests are often referred to collectively as rapid In Vitro Diagnostic (“IVD”) device tests.
Unfortunately, prior rapid IVD test devices are typically useful only for preliminary screening purposes, not as a confirmatory test. Although they can be fast, inexpensive, and simple-to-use, depending on the type of condition being detected, these tests provide a typical accuracy of between 85% and 99%, falling short of the 99.9% or above accuracy generally considered to be necessary for a confirmatory test. To this day, for example, the Western Blot Analytical Assay is the only one reliably used for the confirmatory detection of HIV infection in a clinical laboratory setting worldwide. Due to its multi-step manipulation and verification phases, completion of this type of test can take days, if not weeks. Such a delay can unfortunately lead to further propagation of infectious pathogens such as HIV. Other serious results, such as the metastasis of cancers, can occur while waiting for the results of slower confirmatory tests. There is virtually no generally accepted practical or economical confirmatory rapid diagnostic testing technique for use in a point-of-care setting to rapidly detect serious diseases such as HIV infection and AMI, available in the market place today.
The reasons for the insufficient accuracy in many rapid IVD test devices are primarily due to their current lack of overall higher sensitivity and specificity. Different samples may contain chemicals or particles which inhibit the fluid flow or otherwise interfere with one or both of the affinity binding reactions. Prior devices have attempted to enhance sensitivity or specificity by pretreating various parts of the device with reaction or flow enhancing reagents, pH conditioning chemicals, or even non-specific adhesive blocking molecules which will “block-out” non-analyte molecules which might cause non-specific adhesion, or otherwise compete with the analyte in question for specific binding members, especially on the reaction membrane. These attempts have met with limited success in some types of testing, but do not provide the desired accuracy in many others. Also, pretreatment with two or more of the above pretreatments exacerbates the difficulties in obtaining uniform manufacturing due to potential incompatibilities between the pretreatment chemicals. For example, the pH conditioner might disrupt the effectiveness of the non-specific blocking member molecules. Or, the manufacturing step of pretreating with the second pretreatment chemical can dislodge some of the first pretreatment chemical.
Further, lot-to-lot variation in the manufacture of many IVD test devices can often lead to ambiguous results, such as false negatives as well as weak false positives, so-called “ghost lines” or “phantom lines”. False negatives typically occur when non-specific molecules interfere with the first and/or second affinity binding actions. It has been found that non-analyte molecules can clump together in fluid samples that are not well mixed so that they temporarily prevent access between analytes and binding members. Even temporary interference can prevent an adequate number of labeled analyte complexes and/or ultimately immuno-sandwich complexes from forming. In this way, if a non-analyte molecule or clump of molecules blocks access between analytes and binding members for only a few seconds, it may be enough to induce a false negative result. Further, clumps of non-analyte molecules can carry an overabundance of the labeled mobilizable binding members to the second affinity binding site to generate a false positive.
Chemically non-uniform flows can result in flows having non-uniform first affinity binding by the time they reach the reaction membrane leading to inaccuracies. Such non-uniform flows can be caused by a number of factors. First, some portions of the fluid may flow faster than others from time to time. In those tests having deposits of dried reagent, faster flows tend to reach the dried reagent first. These flows, particularly along the initial fluid front, tend to exhibit a greater degree of first affinity binding per unit fluid or at least uptake of mobilizable labeled binding members, and can potentially carry a greater concentration of clumps of non-analyte molecules which can carry away labeled mobilized binding members. Further, the deposit of dried reagent itself can exhibit portions of higher concentration than others resulting in similar chemical nonuniformity in the flow. Other flows having a lower than average concentration of analyte molecules, and/or having a greater concentration of non-clumped, non-analyte molecules which merely inhibit analyte binding but do not carry away mobilizable labeled binding members, exhibit less apparent first affinity binding. These flow and concentration dis-uniformities are responsible for many of the unsatisfactory results discussed above.
Therefore, there is a need to improve the accuracy of rapid IVD test devices so that Rapid Confirmatory Immunological Testing (“RCIT”) becomes a reality.